CCK-8 kit for 96-well plate cell proliferation How to set up control and sample?

It is recommended to set up at least four types of wells for the 96 well plate proliferation experiment: blank wells (with only culture medium and CCK-8 added, no cells), control group (cells with solvent or culture medium), experimental group (cells with drugs added), and drug background wells (culture medium with drugs and CCK-8 added, no cells). Each group has 5 wells, and it is best to seal the edge wells with PBS or sterile water to reduce evaporation and edge effects. Before adding CCK-8, there is generally no need to change the solution. Simply add 10 microliters of CCK-8 to every 100 microliters of culture medium. When adding the sample, gently tap along the well wall to avoid bubbles. After adding, shake gently and mix well. If the drug itself has color or reducibility, the drug background hole must be made, otherwise the OD will be artificially high or low.

The amount of reagents should be calculated according to your design: for example, adding several background wells to 60 experimental wells, adding 10 microliters of CCK-8 per well, approximately 0.6-0.8 milliliters are needed each time. A 1 milliliter filling is basically only enough for one use, and a 5 milliliter filling can be used multiple times. Whether the price range of 500-800 yuan per box is normal depends on the quantity and specifications. This price range is common in both domestic and imported specifications, but don't just look at the unit price. Consider the cost per hole and whether it comes with instructions and dark storage conditions. CCK-8 should avoid repeated freezing and thawing. It is recommended to store it in the dark at -20 ℃ after packaging, and use it as soon as possible after opening to avoid repeated heating affecting stability.

Serum and phenol red generally have little effect on 450-nanometer readings at conventional concentrations, but if you have a darker medium or noticeable yellowing after cell metabolism, it is best to change fluids or set background wells before adding CCK-8. The difference in color development time between different manufacturers does exist, and the same type of cells may also be different. It is recommended to do a preliminary test first: after adding CCK-8, read it every 0.5-1 hours, find the OD value is still in the linear range and the blank is not high, and the OD of the control group is between 1.2-2.0. Do not fix the time point for an hour. Avoid air bubbles in the holes before reading, which can significantly interfere with OD.